There are, at least, two splicing variants of CaMKII-d, dB and dC, located in nuclear and cytosol compartments, respectively. Our present in vivo and in vitro studies have provided multiple lines of evidence to demonstrate that CaMKII-dB and CaMKII-dC exhibit opposing functional roles in regulating cardiomyocyte viability with CaMKII-dB protective and CaMKII-dC apoptotic. (1) CaMKII Activation Is Required for beta1-AR Induced Apoptosis in Cardiomyocytes and In Vivo. Sustained beta1-AR stimulation markedly increased CaMKII activity in a time-dependent manner in adult mouse cardiomyocytes;this effect was abolished by a specific CaMKII inhibitors AIP or KN93 but not by specific PKA inhibitors Zhu et al.,J. Clin. Invest. 111:617-625, (2003). Importantly, inhibition of CaMKII fully protected myocytes from beta1-AR induced apoptosis. In collaboration with Mark Anderson, we further demonstrated that CaMKII was essential for in vivo apoptotic response to excessive catecholamine stimulation Yang et al., Am. J. Physiol. 291:H3065-H3075, (2006). (2) Increased CaMKII-dC Activity Is Sufficient to Cause Heart Muscle Cell Apoptosis. We specifically enhanced or inhibited CaMKII-dC activity by adenoviral gene transfer of a constitutively active (CA-CaMKII-dC) or a dominant negative CaMKII-dC mutant (DN-CaMKII-dC), respectively. Enforced expression of CA-CaMKII-dC alone caused increased cardiac myocyte apoptosis. The severity of cell apoptosis was closely correlated with CA-CaMKII-dC protein abundance and the kinase activity, suggesting there is a causal relation between activation of CaMKII-dC and cardiomyocyte apoptosis. (3) Activation of Endogenous CaMKII by Various Stimuli That Trigger Myocyte Apoptosis Multiple cell death-inducing stimuli, such as increased intracellular Ca2+ concentration, acidosis, and oxidative stress, increased endogenous CaMKII activity by 2- to 3-fold over baseline. These stimuli markedly trigger myocyte apoptosis in CaMKII-dependent manner. To define the relative contributions of CaMKII-dC versus that of CaMKII-dB to the observed cell death, we suppressed CaMKII-dC activity using the DN-CaMKII-dC, and found that expression of DN-CaMKII-dC inhibited the kinase activity and the associated cell death. These results indicate that activation of CaMKII-dC constitutes a common pathway converging multiple stimuli evoked apoptotic signals in cardiomyocytes (Zhu et al., J. Biol. Chem. 282:10833-10839, 2007). (4) CaMKII-dB Protects Cardiomyocytes Against Apoptosis. In cultured neonatal rat cardiomyocytes, the expression of CaMKII-dB and CaMKII-dC is inversely regulated in response to H2O2-induced oxidative stress with a profound reduction of the former and an increase of the later. In addition, CaMKII-dB expression is remarkably attenuated, while CaMKII-dC expression was augmented at both mRNA and protein levels in rat ischemia/reperfusion (I/R) and myocardium infarction (MI) models. The inhibitory effects of IR and MI on CaMKII-dB are fully prevented by ROS scavengers, indicating ROS oppositely regulates CaMKII-dB and CaMKII-dC expression. Concurrently, MI and H2O2 markedly increase myocyte apoptosis in vivo and in culture, respectively, assayed by Hoechst or TUNEL staining and caspase activation. Most importantly, overexpression of CaMKII-dB using adenoviral gene transfer protects heart cells against oxidative stress-induced apoptosis, assayed by DNA laddering and Hoechst staining (Peng et al., Circ. Res. in press, 2009). (5) Cardioprotection by CaMKII-dB is mediated by phosphorylation of HSF1 and subsequent induction of iHSP70 Using cDNA microarray analysis, real time-PCR and Western blot, we demonstrate that overexpression of CaMKII-dB but not CaMKII-dC elevate expression of heat shock protein 70 (HSP70) family members, including inducible HSP70 (iHSP70) and its homologous (Hst70). Moreover, overexpression of CaMKII-dB leads to phosphorylation and activation of heat shock factor 1 (HSF1), the primary transcription factor responsible for HSP70 gene regulation. Importantly, gene silencing of iHSP70, but not Hst70, abolishes CaMKII-dB-mediated protective effect, indicating that only iHSP70 is required for CaMKII-dB elicited anti-apoptotic signaling. Conclusions: We conclude that, in contrast to CaMKII-dC, CaMKII-dB serves as a potent suppressor of oxidative stress-induced cardiomyocyte apoptosis via phosphorylation of HSF1 and subsequent induction of iHSP70, marking CaMKII-dB as a potential novel therapeutic target of ischemic heart disease. Based on the opposing effects of these cardiac CaMKII isoforms, we envision that a combination of activation of CaMKII-dB with inhibition of CaMKII-dC should be superior to isoform-nonselective inhibition of CaMKII as a potential therapy for the treatment of heart failure or cardiac arrhythmia.